Background: Increasing evidence suggests that high-density lipoprotein (HDL) may play a role in cancer development. We tested the hypothesis that low HDL levels are associated with increased risk of cancer. Methods: Individuals from two population-based cohorts, the Copenhagen General Population Study (2003-2015, N = 107 341), and the Copenhagen City Heart Study (1991-1994, N = 9387) were followed prospectively until end of 2016 to assess low plasma HDL cholesterol and apolipoprotein A1 as risk factors for cancer using Cox proportional hazard regression. Results: During up to 25 years follow-up, we observed 8748 cancers in the Copenhagen General Population Study and 2164 in the Copenhagen City Heart Study. In the Copenhagen General Population Study and compared to individuals with HDL cholesterol ≥ 2.0 mmol/L (≥ 77 mg/dL), multivariable adjusted hazard ratios (HRs) for any cancer were 1.13 (95% confidence interval 1.04-1.22) for individuals with HDL cholesterol of 1.5-1.99 mmol/L (58-77 mg/dL), 1.18 (1.08-1.30) for HDL cholesterol of 1.0-1.49 mmol/L (39-58 mg/dL), and 1.29 (1.12-1.48) for individuals with HDL cholesterol < 1.0 mmol/L (< 39 mg/dL). Correspondingly, compared to individuals with apolipoprotein A1 ≥ 190 mg/dL, HRs for any cancer were 1.06 (0.96-1.17) for individuals with apolipoprotein A1 of 160-189 mg/dL, 1.18 (1.07-1.30) for apolipoprotein A1 of 130-159 mg/dL, and 1.28 (1.13-1.46) for individuals with apolipoprotein A1 < 130 mg/dL. Among 27 cancer types, low HDL cholesterol and/or apolipoprotein A1 were associated with increased risk of multiple myeloma, myeloproliferative neoplasm, non-Hodgkin lymphoma, breast cancer, lung cancer, and nervous system cancer. Results were overall similar in women and men separately, and in the Copenhagen City Heart Study. Conclusions: Low HDL levels were associated with increased risk of several cancers. Increased risk was most pronounced for hematological and nervous system cancer, and to a minor extent for breast and respiratory cancer.